长春新碱通过下调SODD蛋白表达诱导白血病细胞凋亡新机制研究

 目的研究死亡结构域沉默子(silencer of death domains,SODD)、caspase3、caspase8及caspase9在长春新碱诱导Jurkat白血病细胞凋亡过程中的变化,探讨长春新碱诱导肿瘤细胞凋亡的新机制。方法采用Annexin V/PI双标记流式细胞术检测长春新碱(VCR)作用后Jurkat细胞凋亡发生率;采用免疫印迹法分析SODD、caspase3、caspase8及caspase9蛋白表达的变化;ELISA酶联免疫吸附技术检测VCR作用Jurkat细胞后TNF-α分泌的变化;采用RT-PCR检测VCR对细胞,TNFR1mRNA表达的调节。结果Jurkat白血病细胞SODD蛋白高表达且高表达的SODD蛋白抑制肿瘤细胞凋亡,VCR能特异下调SODD蛋白的表达,有效诱导Jurkat细胞凋亡,但并不影响细胞TNF-α的分泌及TNFR1的表达;VCR诱导细胞凋亡过程中caspase3、caspase8酶原呈时间依赖性逐渐被水解剪切,而caspase9在该凋亡过程中无明显变化趋势。结论VCR下调SODD蛋白表达并启动外源性凋亡途径caspases级联(caspase8、caspase3),最终诱导Jurkat细胞凋亡,且VCR下调SODD蛋白的表达无需激活TNF/TNFR1信号途径即可导致凋亡的发生。

New Mechanism of Jurkat T Cells Apoptosis Induced by Vincristine Via Inhibiting the Expression of SODD Protein

OBJECTIVE To study the changes of silencer of death domains(SODD),caspase3,caspase8 and caspase9 during the apoptotic process of human acute lymphoblastic leukemia Jurkat T cells induced by VCR and explore the new mechanism of tumor cell apoptosis induced by VCR.METHODS After Jurkat cells were induced by VCR,translocated phosphatidylserine was labeled with Annexin V/PI.The apoptosis incidence was measured by FCM.The changes of the expression of SODD,caspase3,caspase8 and caspase9 proteins were determined by western blotting.The secretion of TNF-α was determined by ELISA.The changes of TNFR1 mRNA were determined by RT-PCR.RESULTS Over-expression of SODD in Jurkat cells inhibited the apoptosis of tumor cell,VCR induced the apoptosis of Jurkat cells via down-regulating SODD.caspase3 and caspase8 were hydrolysated in a time-dependent manner,in the process,caspase9 was not been deregulated.While the secretion of TNF-α and the expression of TNFR1 in Jurkat cells were not changed by VCR.CONCLUSION VCR Induces the apoptosis of Jurkat cells via downregulating protein expression of SODD which need not activate the signal pathway of TNF/TNFR1 and starts up extrinsic apoptosis pathway namely cascades including caspase8 and caspase3.